An estimated 300 million people worldwide are infected with hepatitis C virus (HCV) and a significant percentage of infected individuals develop cirrhosis, hepatic failure and hepatocellular carcinoma (HCC). This results in approximately 8-10,000 deaths annually among the 3.9 million infected individuals in the United States. HCV infection, thus, has emerged as the leading cause of cirrhosis in the United States and for patients with established cirrhosis related to HCV infection, the 5-year risk of liver failure is approximately 18% and that of HCC is approximately 7%. Therefore, HCV-related liver failure had become the most frequent indication for orthotopic liver transplantation (OLT). Therefore, a better understanding of the immune mediated mechanisms underlying the pathogenesis of chronic HCV infection, specifically in immunocompromised HCV-infected OLT recipients is urgently needed. The specific goals of this proposal are: 1) To define HCV-specific CD4+ and CD8+ T cell immune responses in HCV-infected OLT recipients in order to test the hypothesis that the lack of HCV-related allograft injury is associated with the development of anti-HCV CD4+ and CD8+ T cell responses. HCV specific CD4+ and CD8+ T cell responses will be assessed both in the peripheral circulation and in the graft infiltrating cells by means of granzyme B, IFN-n, and IL-5 ELISPOT analyses. 2) To determine whether immune functional assays for detection of alloreactivity and anti-HCV reactivity can differentiate between HCV recurrence and acute cellular rejection, since this is one of the main problems existing today in the management of the HCV-infected OLT recipient. Towards this, the CD4+ and CD8+ T cell alloreactivity and/or HCV-specific reactivity will be determined both in peripheral blood as well as in graft-infiltration lymphocytes at the time of suspected rejection by means of granzyme B, IFN-E], and IL-5 ELISPOT assays. 3) To define HCV-specific CD8+ T cell immune responses in patients with HCV-related HCC in order to test the hypothesis that HCC-positive patients will display a different profile of HCV-specific CD8+ T cell reactivity as compared to HCV-infected HCC-negative patients. These studies will also identify the HCV-derived peptides expressed on HCC cells and their most common mutations. The overall goal of this proposal is to better define the immune mediated mechanisms underlying the pathogenesis of chronic HCV infection in immunocompromised OLT recipients and HCC patients which will aid in designing new strategies for treatment of these diseases. Further, this project is intended to develop an in vitro method for differentiating HCV recurrence and acute cellular rejection following OLT that will have immediate benefit in the management of the HCV-infected OLT recipients. [unreadable] [unreadable]